The genetic background for the onset of cancer is alterations in proto-oncogenes, oncogenes and tumour suppressor genes. Proto-oncogenes are normal genes of the cell which have the potential of becoming oncogenes. All oncogenes code for and function through a protein. In the majority of cases they have been shown to be components of signal transduction pathways. Oncogenes arise in nature from proto-oncogenes through point mutations or translocations, thereby resulting in a transformed state of the cell harbouring the mutation. Cancer develops through a multi-step process involving several mutational events in oncogenes and tumour suppressor cells
In its simplest form, a single base substitution in a proto-oncogene may cause the encoded protein to differ in one amino acid.
In experimental models involving murine tumours, it has been shown that point mutations in intracellular “self”-proteins may give rise to tumour rejection antigens consisting of peptides differing in a single amino acid from the normal peptide. The T cells recognizing these peptides in the context of major histocompatibility (MHC) molecules on the surface of the tumour cells are capable of killing the tumour cells and thus rejecting the tumour from the host. (Boon, T. et al, Cell 1989, Vol. 58, p. 293-303)
In the last three decades, particular effort has been devoted to the analysis of antibodies to human tumour antigens. It has been suggested that such antibodies could be used both for diagnostic and therapeutic purposes, for instance in connection with an anti-cancer agent. One problem is that antibodies can only bind to tumour antigens that are exposed on the surface of tumour cells. For this reason the efforts to produce a cancer treatment based on the immune system of the body has been less successful than expected.
Antibodies typically recognise free antigens in native conformation and can potentially recognise almost any site exposed on the antigen surface. In contrast to the antibodies produced by B cells, T cells recognise antigens only in the context of MHC molecules, designated HLA (human leukocyte antigen) in humans, and only after appropriate antigen processing, usually consisting of proteolytic fragmentation of the protein, resulting in peptides that fit into the groove of the MHC molecules. This enables T cells to recognise peptides derived from intracellular proteins. T cells can thus recognise aberrant peptides derived from anywhere in the tumour cell, when displayed on the surface of the tumour cell by MHC molecules. The T cell can subsequently be activated to eliminate the tumour cell harbouring the aberrant peptide.
T cells may control the development and growth of cancer by a variety of mechanisms. Cytotoxic T cells, both HLA class I restricted CD8+ and HLA Class II restricted CD4+, may directly kill tumour cells carrying the appropriate tumour antigens. CD4+ helper T cells are needed for induction and maintenance of cytotoxic T cell responses as well as for antibody responses, and for inducing macrophage and lymphokine-activated killer cell (LAK cell) killing.
Many oncogenes and their protein products have been identified. In addition, it has been shown that the T cell repertoire of a healthy person includes T cells with specificity against a synthetic peptide fragment derived from one p21 RAS oncogene product, when presented on an appropriate HLA molecule. Furthermore, it is anticipated that approximately 20% of all cancers are associated with a mutation in the RAS oncogene.
WO 92/14756 discloses synthetic peptides and fragments of oncogene protein products which elicit T cell immunity, for use in vaccines against cancers associated with RAS and compositions for the treatment of cancer. The peptides correspond to an active fragment of the oncogene as presented by the cancer cell and include a mutation in one or more positions corresponding to the oncogene mutation. This document discloses mutations at positions 12, 13 and 61 of the RAS protein and specifically discloses G12A, G12V, G12C, G12S, G12K, G12D, G12R, Q61R, Q61K, Q61L, Q61H, G13V and G13D mutations. While this document mentions that vaccines may comprise a selection of peptides having the most common mutations found in oncogene proteins, it does not suggest any specific combinations of peptides.
WO 00/66153 discusses synthetic peptide mixtures which elicit T cell immunity for use in cancer vaccines. The peptide mixtures consist of RAS p21 mutant peptides and this document specifically discloses G12A, G12C, G12D, G12R, G12S, G12V, Q61H, Q61K, Q61L, Q61R and G13D mutations. This document also discloses that the immune response elicited by a cocktail of peptides was significantly higher than that elicited by a single peptide. However, it does not suggest the use of a peptide vaccine in a combined treatment of cancer with any other form of therapy.
Gjertsen et al. (Int. J. Cancer 2001, 92, p. 441-450) discloses a phase I/II trial involving patients with adenocarcinoma of the pancreas vaccinated with synthetic mutant RAS peptides in combination with granulocyte-macrophage colony-stimulating factor. This trial used single peptide vaccines or a mixture of four mutant peptides. The combination vaccine consisted of the four most common K-RAS mutations found in pancreatic adenocarcinoma, namely peptides having a G12V, a G12D, a G12C or a G12R mutation. This document does not suggest that combination therapy with anti-metabolite chemotherapeutic agents may be effective.
Wedén et al. (Int. J. Cancer 2010, 128(5), p. 1120-1128) reports the long-term follow-up of patients with pancreatic adenocarcinoma vaccinated with synthetic mutant RAS peptides. The vaccine consisted of either a single RAS peptide or a cocktail of seven RAS peptides. In particular, the seven RAS peptides used in this trial had a G12A, a G12C, a G12D, a G12R, a G12S, a G12V or a G13D mutation. There is no mention of a combination therapy with anti-metabolite chemotherapeutic agents.
Prior et al. (Cancer Res. 2012, 72(10), p. 2457-2467) discloses that different types of cancer are coupled to mutation of a particular RAS isoform and that each isoform has a distinctive codon mutation signature. In addition, Prior et al. discloses that a total of 18 mutations occur in positions 12, 13 and 61 of the RAS protein, with six mutations occurring in each position. This review also discusses the effects of these mutations on RAS function and the potential mechanisms leading to differential patterns of RAS isoform mutations.
A previously unrelated treatment of cancer patients has been systemic administration of the chemotherapeutic agent gemcitabine. Gemcitabine is currently a frequent chemotherapeutic treatment for patients with various cancers, such as non-small cell lung cancer, pancreatic cancer, bladder cancer and breast cancer. In particular, gemcitabine is frequently used to treat advanced pancreatic cancer.
Gemcitabine is an anti-metabolite chemotherapeutic agent, specifically a pyrimidine analogue. The triphosphate analogue of gemcitabine replaces cytidine during DNA replication and incorporation into the elongating DNA strand halts further DNA synthesis after addition of one more nucleotide as the DNA polymerase is unable to proceed. Further mechanisms of action are thought to include inhibition of ribonucleoside reductase, leading to a depletion of deoxyribonucleotide pools necessary for DNA synthesis, and competition with deoxycytidine triphosphate as an inhibitor of DNA polymerase. These actions result in necrosis, leading to an arrest of tumour growth. This action also means that anti-metabolite chemotherapeutic agents are toxic to other actively dividing cells such as T cells. The preferred regimen involves using chemotherapy first, followed by vaccination to enhance later induction of immune responses from killed cancer cells.
While some drug combinations, such as FOLFIRINOX, have been shown to be more effective than gemcitabine in patients with metastatic pancreatic adenocarcinoma, therapy with this combination was associated with a high incidence of side effects (Conroy et al., N Engl J Med 2011, vol. 364(19), P. 1817-1825). It is desirable, therefore, to find novel strategies for the treatment of cancer.
Oettle et al. (JAMA 2007, 297(3), p. 267-277) reports an investigation of the use of gemcitabine as adjuvant chemotherapy in resectable pancreatic cancer, and discloses that the effect of gemcitabine on disease-free survival was significant in patients with either R0 or R1 resection. In particular, this study discloses that postoperative gemcitabine significantly delayed the development of recurrent disease after complete resection of pancreatic cancer, compared to patients to whom gemcitabine was not administered following resection. However, this study does not suggest that gemcitabine could be useful in combination with other pharmaceutical treatments of cancer.
Bauer et al. (Cancer Immunol. Immunother. 2014, 63, p. 321-333), discloses that gemcitabine has a negative influence on dendritic cell (DC) vaccine-induced T-cell proliferation. In particular, this study showed that delayed administration of gemcitabine, as compared to the DC vaccine, did not improve patient's immune response, while concomitant administration of gemcitabine and the DC vaccine significantly impaired the vaccine-induced immune response.
Thus, there is a need to provide further and more effective cancer treatments.